DCM Fraction Research Articles

Anti-invasive Activity against Cancer Cells of Phytochemicals in Red Jasmine Rice (Oryza sativa L.)

Red rice contains pharmacological substances including phenolics, oryzanol, tocotrienol and tocopherol. Recently, red rice extract has been employed as a source of antioxidants for inhibition of tumor growth. This study was carried out to evaluate the anti-invasion effects of red rice extract fractions on cancer cells. It was found that at 100 μg/ml of crude ethanolic extract (CEE), hexane fraction (Hex) and dichloromethane fraction (DCM) could reduce HT1080 and MDA-MB-231 cancer cell invasion. Hex and DCM revealed higher potency levels than CEE, whereas an ethyl acetate fraction (EtOAc) had no effect. Gelatin zymography revealed that Hex decreased the secretion and activity of matrix metalloproteinase-2 and -9 (MMP-2 and-9). In contrast, the DCM fraction exhibited slightly effect on MMPs secretion and had no effect on MMPs activity. Collagenase activity was significantly inhibited by the Hex and DCM fractions. High amounts of γ-oryzanol and γ-tocotrienol were found in the Hex and DCM fractions and demonstrated an anti-invasion property. On the other hand, proanthocyanidin was detected only in the CEE fraction and reduced MDA-MB-231 cells invasion property. These observations suggest that proanthocyanidin, γ-oryzanol and γ-tocotrienol in the red rice fractions might be responsible for the anti invasion activity. The red rice extract may have a potential to serve as a food-derived chemotherapeutic agent for cancer patients.

Antioxidant activities and phenolic contents of three red seaweeds (Division: Rhodophyta) harvested from the Gulf of Mannar of Peninsular India.

The antioxidant activities of methanol extract and its solvent fractions (n-hexane, dichloromethane and ethyl acetate) of three red seaweeds (Hypnea musciformis, H. valentiae, and Jania rubens) collected from the Gulf of Mannar of South eastern coast of India were evaluated, using different in vitro systems, viz., DPPH, ABTS, HO radical scavenging activities, H2O2 scavenging ability, Fe(2+) ion chelating ability and reducing potential. Folin-Ciocalteu method was used to determine the total phenolic content of the extracts/fractions, and the results were expressed as mg of gallic acid equivalent (GAE)/g of the seaweed extracts/fractions. Thiobarbituric acid-reactive substances (TBARS) inhibition assay was employed to assess the ability of the seaweed extracts/fractions to inhibit lipid oxidation. Ethyl acetate (EtOAc) fractions of H. musciformis exhibited significantly higher total phenolic content (205.5mg GAE/g), DPPH· scavenging activity (IC50 0.6mg/mL), ABTS(.+) scavenging activity (IC50 0.51μg/mL), Fe(2+) chelating ability (IC50 0.70mg/mL), H2O2 scavenging activity (IC50 0.39mg/mL), reducing ability (Abs700 nm 1.46) and lipid peroxidation inhibitory ability (2.71 MDAEC/kg) (P < 0.05) compared to its n-hexane, DCM fractions, crude MeOH extract and MeOH extracts/fractions of H. valentiae and J. rubens. DCM fraction of J. rubens showed significantly higher hydroxyl radical scavenging activity (IC50 0.55mg/mL) compared with H. musciformis and H. valentiae (P < 0.05). This study indicated the potential use of red seaweeds, in particular, H. musciformis as candidate species to be used as food supplement for increasing the shelf-life of food industry, and candidates in combating carcinogenesis and inflammatory diseases.

Dopaminergic effect of Alstonia scholaris Linn. R.Br. in experimentally induced anxiety

Aim of the studyThe anti-anxiety activity of the ethanolic extract of leaves of Alstonia scholaris (Family: Apocynaceae) has been well established. The aim of the present study is to evaluate the anti-anxiety activity of the dichloromethane fraction (known as DCM) fractionated from ethanolic extract of leaves of A. scholaris (known as EEAS). Materials and methodsDCM fraction (10 and 30mg/kg) was tested against various anti-anxiety models viz. elevated plus maze, open field, hole board, light dark test, mirror chamber and foot shock induced aggression models. The false positive results were overruled by actophotometer and rotarod tests. The change in brain monoamines was estimated. The possible dopaminergic effect was tested by haloperidol-induced catalepsy and 6-hydroxy dopamine induced Parkinsonism. ResultsThe DCM fraction of EEAS was found to be significantly active in all the tested models of anxiety, which confirmed their anti-anxiety activity. Lack of significant change in locomotion test indicated that DCM did not have stimulant or sedative effects. There was also no significant change in the time spent in the rotarod, which indicated its lack of motor or muscle in coordination. Increase in brain dopamine levels and reversal of dopamine-induced catalepsy confirmed that the mechanism of anti-anxiety activity is mediated through dopamine. The effect of DCM in 6-hydroxy dopamine-induced anxiety was conclusive of its dopaminergic activity. ConclusionThe present study suggests that DCM possess anti-anxiety and the apparent mechanism of action is through dopaminergic mediation.

In vitro anti-cancer and anti-oxidant activity of different fractions of Diospyros peregrina unripe fruit extract

Background: Diospyros peregrina (Ebenaceae) is a medium-sized evergreen tree having ethnomedicinal significance as an aphrodisiac, astringent, bactericide and for the treatment of diarrhoea, cholera, dysentery, fever, malaria and diabetes. Objectives: To evaluate the anti-oxidant and anti-cancer activity of different extracts and fractions of matured unripe fruits of Diospyros peregrina on various in vitro models. Materials and Methods: Anti-oxidant and free radical scavenging activity of various extracts and fractions were determined by DPPH, ABTS and Alkaline DMSO methods. In vitro cytotoxicity of different extracts and fractions of matured unripe fruits of Diospyros peregrina was evaluated on MCF-7 and Hep-G2 cell lines by MTT and SRB methods. Results: Among the extracts and fractions studied for antioxidant activity n-butanol fraction (NBF) and aqueous extract (AE) showed potential scavenging effect against DPPH, ABTS and superoxide free radicals. In vitro cytotoxicity assays of extracts and fractions by MTT and SRB methods revealed that dichloromethane fraction (DCMF) exhibited maximum cytotoxicity on both MCF-7 and Hep G2 cell lines. Conclusion: The higher anti-oxidant potential of n-butanol fraction (NBF) and aqueous extract (AE) may be due to the presence of phenolic compounds, flavonoids or flavonoid glycosides. Dichloromethane fraction (DCMF) exhibited maximum cytotoxicity on both MCF-7 and Hep G2 cell lines. The cytotoxic activity of DCM fraction may be attributed due the presence of triterpenoids.

Abstract B170: Identification of compounds as cytotoxic agents and inhibitors of protein kinase C, a key enzyme responsible for the control, growth, division, and differentiation of cells.

Abstract Cancer is the second leading cause of death worldwide in humans. Present treatment options available are surgery, radiation and chemotherapy. However, all treatment procedures suffer from serious side effects and toxicity. Hence there is an urgent need to develop alternative medicines and/or drug candidates which act specifically on the cancer cells. Protein kinases regulate a multitude of physiological processes by the transfer of phosphate residues and there by playing a central role in control of growth, division and differentiation of cells. There is sufficient evidence that over expression or elevated activity or loss of activity control of different protein kinases is involved in initiation and progression of tumors. Protein kinase C (PKCs) with multiple functions is an interesting target for cancer therapy. PKCs are also known to modulate multidrug resistance, and thus the use of PKC modulators in combination with classical cytotoxic drugs could be beneficial. Experimental procedure: In a joint effort with the local Amchis and scientists of IIIM, Jammu, seven medicinal plants were procured namely Christolia crassifolia, Tanacetum gracile, Echinecea purpurea (root and aerial part), Perovskia abrotanoides Rhodiola imbricata, Saussurea bracteata and Actinocarya tibetica. The plant materials were extracted by using a mixture of organic solvents (MeOH: water, 9:1). The extract was then concentrated below 50°C under reduced pressure. The concentrated methanol extract was again fractionated with n-hexane, dichloromethane, ethylacetate and aqueous fractions respectively. Solvent was evaporated under reduced pressure from each fraction. Each fraction and extracts after complete removal of solvent were evaluated for cytotoxic activity by MTT based colorimetric assay. Separation of fractions from the concentrated extract of the plant (Tanacetum gracile) was done by column chromatography using n-hexane as mobile phase and then elution was carried out in n-hexane and ethyl acetate with solvent gradient. The fractions were collected and pooled on the basis of TLC pattern. Six compounds have been isolated and their structural analysis by various spectroscopic techniques like NMR, UV, IR and Mass spectroscopy is in process. All these compounds will be tested for protein kinase enzyme inhibition activity. This could provide a lead in further optimization of activity for use in drug development. Results: Four plants; hexane and DCM fraction of Tanacetum gracile, hexane and DCM fraction of Echinecea purpurea (root and aerial part), hexane and DCM fraction of Perovskia abrotanoides and hexane, DCM and EtOAc fractions of Saussurea bracteata showed substantial cytotoxic activity. On an average the IC50 values of these components were between 6 to 60μg/mL. Cytotoxicity of these six purified compounds was evaluated on PC-3 and HeLa cells by MTT Assay. Conclusions: The results in the present study clearly demonstrates that T.gracile, P.abrotanoides, S.bracteata and E.purpurea extracts could exhibit an antiproliferative effect on human prostate cancer and tumor cells. These results reveal the initial potential of these medicinal plants as cytotoxic agents for therapy against human prostate cancer and tumor. It is therefore worthwhile to isolate the active compounds and identity their structures. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2011 Nov 12-16; San Francisco, CA. Philadelphia (PA): AACR; Mol Cancer Ther 2011;10(11 Suppl):Abstract nr B170.

Bioassay-guided isolation of norviburtinal from the root of Rehmannia glutinosa, exhibited angiogenesis effect in zebrafish embryo model

Ethnopharmacological relevanceThe root of Rehmannia glutinosa (RR) is commonly used as a wound-healing agent in various traditional Chinese herbal formulae; while angiogenesis is one of the crucial aspects in wound-healing. Aim of the studyThe objective of the present study was to investigate the angiogenesis effects of RR aqueous crude extract and its active component(s) using zebrafish model. Materials and methodsThe in vivo angiogenesis effect was studied using transgenic TG(fli1:EGFP)y1/+(AB) zebrafish embryos by observing the capillary sprouts formation in sub-intestinal vessel (SIV) of zebrafish embryos after 72h post-fertilization under fluorescence microscopy. ResultsOur results indicated that RR aqueous crude extract (250μg/ml) exhibited significant angiogenesis effect, with an increase in capillary sprouts formation in SIV. Following sequential solvent partition of the RR aqueous crude extract with dichloromethane, ethyl acetate and n-butanol successively, the dichloromethane fraction (DCM) was found to have the most sprouts formation in the SIV region. Subjected to column chromatography, DCM fraction was further fractionated into six sub-fractions and among these tested, the sub-fraction C2 exhibited the most potent angiogenesis effect. The major component, C2A, was isolated and identified as norviburtinal using nuclear magnetic resonance (NMR) and mass spectrometry (MS). The compound norviburtinal (at 50μg/ml) was shown to possess significant angiogenesis effect in zebrafish model (p<0.001). ConclusionsNorviburtinal was, for the first time, found in the extract of RR and possessed novel angiogenesis effect. Bioassay-guided fractionation suggested that norviburtinal was not the only active component responsible for the angiogenesis effect of RR.

Bioassay-directed fractionation and salmonella mutagenicity of automobile and forklift diesel exhaust particles.

Many pulmonary toxicity studies of diesel exhaust particles (DEPs) have used an automobile-generated sample (A-DEPs) whose mutagenicity has not been reported. In contrast, many mutagenicity studies of DEPs have used a forklift-generated sample (SRM 2975) that has been evaluated in only a few pulmonary toxicity studies. Therefore, we evaluated the mutagenicity of both DEPs in Salmonella coupled to a bioassay-directed fractionation. The percentage of extractable organic material (EOM) was 26.3% for A-DEPs and 2% for SRM 2975. Most of the A-EOM (~55%) eluted in the hexane fraction, reflecting the presence of alkanes and alkenes, typical of uncombusted fuel. In contrast, most of the SRM 2975 EOM (~58%) eluted in the polar methanol fraction, indicative of oxygenated and/or nitrated organics derived from combustion. Most of the direct-acting, base-substitution activity of the A-EOM eluted in the hexane/dichloromethane (DCM) fraction, but this activity eluted in the polar methanol fraction for the SRM 2975 EOM. The direct-acting frameshift mutagenicity eluted across fractions of A-EOM, whereas > 80% eluted only in the DCM fraction of SRM 2975 EOM. The A-DEPs were more mutagenic than SRM 2975 per mass of particle, having 227 times more polycyclic aromatic hydrocarbon-type and 8-45 more nitroarene-type mutagenic activity. These differences were associated with the different conditions under which the two DEP samples were generated and collected. A comprehensive understanding of the mechanisms responsible for the health effects of DEPs requires the evaluation of DEP standards for a variety of end points, and our results highlight the need for multidisciplinary studies on a variety of representative samples of DEPs.